Low background catalytic redox recycling coupled with hybridization chain reaction amplification for highly sensitive electrochemical aptamer luteinizing hormone assay.

The concentration variation of luteinizing hormone (LH) regulates the cell cycle of oocyte meiosis and significantly affect the whole reproductive cycle. Sensitively quantifying the LH biomarker therefore plays an important role for reproductive disease diagnosis. By coupling a new low background catalytic redox recycling strategy with hybridization chain reaction (HCR), we propose a highly sensitive bio-electrochemical aptamer LH sensing method.

Target-responsive triplex aptamer nanoswitch enables label-free and ultrasensitive detection of antibody in human serum via lighting-up RNA aptamer transcriptions.

Accurate and sensitive monitoring of the concentration change of anti-digoxigenin (Anti-Dig) antibody is of great importance for diagnosing infectious and immunological diseases. Combining a novel triplex aptamer nanoswitch and the high signal-to-noise ratio of lighting-up RNA aptamer signal amplification, a label-free and ultrasensitive fluorescent sensing approach for detecting Anti-Dig antibodies is described. The target Anti-Dig antibodies recognize and bind with the nanoswitch to open its triplex helix stem structure to release Taq DNA polymerase and short ssDNA primer simultaneously, which activates the Taq DNA polymerase to initiate downstream strand extension of ssDNA primer to yield specific dsDNA containing RNA promoter sequence.

Target-promoted autocatalytic hairpin assembly of bivalent DNAzymes for sensitive and label-free electrochemical metallothionein assay.

Metallothionein (MT) has shown to be an important biomarker for environmental monitoring and various diseases, due to its significant binding ability to heavy metal ions. On the basis of such a characteristic and the Hg-stabilized DNA duplex (Hg-dsDNA) probe, as well as a new autocatalytic hairpin assembly (aCHA)/DNAzyme cascaded signal enhancement strategy, the construction of a highly sensitive and label-free electrochemical MT biosensor is described. Target MT molecules bind Hg in Hg-dsDNA to disrupt the duplex structure and to release ssDNA sequences, which trigger subsequent aCHA for efficient production of mimic aCHA triggering strands and many bivalent DNAzymes.

Simultaneous and Sensitive Sensing of Intracellular MicroRNA and mRNA for the Detection of the PI3K/AKT Signaling Pathway in Live Cells.

Simultaneous detection of the concentration variations of microRNA-221 (miRNA-221) and PTEN mRNA molecules in the PI3K/AKT signaling pathway is of significance to elucidate cancer cell migration and invasion, which is useful for cancer diagnosis and therapy. In this work, we show the biodegradable MnO nanosheet-assisted and target-triggered DNAzyme recycling signal amplification cascaded approach for the specific detection of the PI3K/AKT signaling pathway in live cells via simultaneous and sensitive monitoring of the variation of intracellular miRNA-221 and PTEN mRNA. Our nanoprobes enable highly sensitive and multiplexed sensing of miRNA-221 and PTEN mRNA with low detection limits of 23.

Hg-triggered cascade strand displacement assisted CRISPR-Cas12a for Hg quantitative detection using a portable glucose meter.

CRISPR-Cas12a is a powerful and programmable tool that has revolutionized the field of biosensing. However, the construction of a CRISPR-Cas12a-mediated portable system for on-site and quantitative detection of mercury ion (Hg) has yet to be explored. By integrating a target-triggered cascade toehold-mediated strand displacement reaction (TSDR) and CRISPR-Cas12a, we herein construct a portable on-site biosensor for the quantitative, sensitive, and selective detection of Hg with a glucose meter.

Targeted Delivery of DNA Framework-Encapsulated Native Therapeutic Protein into Cancer Cells.

A protein-based therapy is significantly challenged by the successful delivery of native proteins into the targeted cancer cells. We address this challenge here using an all-sealed divalent aptamer tetrahedral DNA framework (asdTDF) delivery platform, in which the protein drug is encapsulated inside the cavity of the framework stoichiometrically a reversible chemical bond. The ligase-assisted seal of the nicks results in highly enhanced TDF stability of the against nuclease digestion to effectively protect the therapeutic protein from degradation.

Lighting-up RNA aptamer transcription synchronization amplification for ultrasensitive and label-free imaging of microRNA in single cells.

Sensitive imaging of intracellular microRNAs (miRNAs) in cells is of great significance in clinical diagnoses and disease treatments, and it remains a major challenge to achieve this goal. Herein, we report a new in situ rolling circle transcription synchronization machinery (RCTsm) of lighting-up RNA aptamer strategy for highly sensitive imaging and selective differentiation of miRNA expression levels in cells. Such a RCTsm approach utilizes a DNA promoter to recycle the target miRNAs to trigger the initiation of multiple RCT process for the yield of many lighting-up RNA aptamers.

Efficient and Exponential Rolling Circle Amplification Molecular Network Leads to Ultrasensitive and Label-Free Detection of MicroRNA.

MicroRNAs (miRNAs) are useful biomarkers for the diagnosis of a variety of cancers. However, it is a major challenge to detect miRNAs, considering their high sequence similarity, low concentration, and small size nature. With the establishment of an efficient rolling circle amplification (RCA) molecular network by target-driven polymerization/nicking reactions, we present here an exponential amplification strategy for detecting miRNA in a label-free way with ultrahigh sensitivity.

Target-mediated base-mismatch initiation of a non-enzymatic signal amplification network for highly sensitive sensing of Hg.

Because of its adverse environmental effects, the establishment of convenient methods for monitoring Hg2+ with ultrahigh sensitivity is important to human health. With a new target-mediated base-mismatch initiation of a signal amplification network strategy, we describe the development of a simple fluorescence sensing approach for detecting Hg2+ in water samples with high sensitivity. The assistant DNA probes trigger the catalytic hairpin assembly (CHA) of two elaborately designed hairpins for the formation of many Mg2+-dependent DNAzymes via T-Hg2+-T base mismatch hybridization.

Cascaded multiple recycling amplifications for aptamer-based ultrasensitive fluorescence detection of protein biomarkers.

Highly sensitive detection of molecular biomarkers plays a significant role in diagnosing various types of diseases at the early stage. We demonstrated in this paper an ultrasensitive aptamer-based fluorescence method for detecting mucin 1 (MUC1) in human serum via a cascaded multiple recycling signal amplification strategy. The MUC1 target molecules present in the samples cause structure switching of the hairpin aptamer probes, which initiates three cascaded recycling cycles for the cleavage of the fluorescently quenched signal probes to recover significant fluorescence for highly sensitive detection of MUC1.

Netlike hybridization chain reaction assembly of DNA nanostructures enables exceptional signal amplification for sensing trace cytokines.

The monitoring and detection of molecular biomarkers play crucial roles in disease diagnosis and treatment. In this work, we proposed a target-responsive netlike hybridization chain reaction (nHCR) DNA nanostructure construction method, which can offer an exceptional signal enhancement, for highly sensitive fluorescence detection of cytokine, interferon-gamma (IFN-γ). The presence of the target cytokine can lead to the conformational change of the aptamer recognition hairpin probes and the liberation of the nHCR initiator strands, which further trigger the nHCR process between two dye-labeled and double hairpin-structured probes to form netlike DNA nanostructures.

Hairpin/DNA ring ternary probes for highly sensitive detection and selective discrimination of microRNA among family members.

The detection and quantification of microRNA (miRNA) plays essential roles in clinical and biomedical research. Yet, it is of major challenge to sense miRNA with high degree of selectivity and sensitivity due to its unique characteristics of short length, similarity of sequence among family members and low abundance. Here, with the design of a new hairpin/DNA ring ternary probe, we describe the development of a rolling circle amplification (RCA) method for sensitively and selectively sensing miRNA from cancer cells.

Programmable DNA Ring/Hairpin-Constrained Structure Enables Ligation-Free Rolling Circle Amplification for Imaging mRNAs in Single Cells.

Self-assembled functional DNA structures have proven to be excellent materials for designing and implementing a variety of nanoscale devices. We demonstrate here that a rationally designed and programmable DNA ring/hairpin-constrained structure can achieve in situ ligation-free rolling circle amplification (RCA), which further leads to highly specific, sensitive, and multicolor imaging of mRNA molecules in single cells. Such a structure aims at addressing current challenges in terms of simplicity, sensitivity, and multiplexing capability related to the detection and imaging of intracellular mRNA sequences.

Silver ion-stabilized DNA triplexes for completely enzyme-free and sensitive fluorescence detection of transcription factors via catalytic hairpin assembly amplification.

Transcription factors play important roles in gene regulation and have been identified as promising biomarkers for disease diagnosis. On the basis of a new Ag-stabilized DNA triplex probe and catalytic hairpin assembly (CHA) signal amplification, we have established a completely enzyme-free and sensitive method for simple fluorescence detection of NF-κB p50 (nuclear factor-kappa B), a transcription factor. We found that the employment of Ag to stabilize the DNA triplex structure could effectively reduce the background noise.

Bio-cleavable nanoprobes for target-triggered catalytic hairpin assembly amplification detection of microRNAs in live cancer cells.

The monitoring and imaging of intracellular microRNAs (miRNAs) with specific sequences plays a vital role in cell biology as it can potentially elucidate many cellular processes and diseases related to miRNAs in living cells with accurate information. However, the detection of trace amounts of under-expressed intracellular miRNAs in living cells represents one of the current major challenges. In an effort to address this issue, we describe the establishment of an in cell catalytic hairpin assembly (CHA) signal amplification strategy for imaging under-expressed intracellular miRNAs in this work.

A target-responsive autonomous aptamer machine biosensor for enzyme-free and sensitive detection of protein biomarkers.

Highly sensitive and selective detection of protein disease markers is crucial for fundamental research and disease diagnosis. In this work, based on a new target-triggered autonomous aptamer machinery amplification approach, we developed a simple and sensitive sensing platform for the detection of a cancer biomarker, platelet-derived growth factor BB (PDGF-BB), in human sera. The target PDGF-BB binds the fluorescently quenched hairpin aptamer probes and causes structural and conformational changes in the aptamers to recover their fluorescence.

A catalytic and dual recycling amplification ATP sensor based on target-driven allosteric structure switching of aptamer beacons.

Abnormal concentrations of ATP are associated with many diseases and cancers, and quantitative detection of ATP is thus of great importance for disease diagnosis and prognosis. In the present work, we report a new dual recycling amplification sensor integrated with catalytic hairpin assembly (CHA) to achieve high sensitivity for fluorescent detection of ATP. The association of the target ATP with the aptamer beacons causes the allosteric structure switching of the aptamer beacons to expose the toehold regions, which hybridize with and unfold the fluorescently quenched hairpin signal probes (HP1) to recycle the target ATP and to trigger CHA between HP1 and the secondary hairpin probes (HP2) to form HP1/HP2 duplexes.

Click chemistry-mediated cyclic cleavage of metal ion-dependent DNAzymes for amplified and colorimetric detection of human serum copper (II).

The determination of the level of Cu plays important roles in disease diagnosis and environmental monitoring. By coupling Cu-catalyzed click chemistry and metal ion-dependent DNAzyme cyclic amplification, we have developed a convenient and sensitive colorimetric sensing method for the detection of Cu in human serums. The target Cu can be reduced by ascorbate to form Cu, which catalyzes the azide-alkyne cycloaddition between the azide- and alkyne-modified DNAs to form Mg-dependent DNAzymes.

A DNA-Fueled and Catalytic Molecule Machine Lights Up Trace Under-Expressed MicroRNAs in Living Cells.

The detection of specific intracellular microRNAs (miRNAs) in living cells can potentially provide insight into the causal mechanism of cancer metastasis and invasion. However, because of the characteristic nature of miRNAs in terms of small sizes, low abundance, and similarity among family members, it is a great challenge to monitor miRNAs in living cells, especially those with much lower expression levels. In this work, we describe the establishment of a DNA-fueled and catalytic molecule machinery in cell signal amplification approach for monitoring trace and under-expressed miRNAs in living cells.

Biodegradable MnO Nanosheet-Mediated Signal Amplification in Living Cells Enables Sensitive Detection of Down-Regulated Intracellular MicroRNA.

The monitoring of intracellular microRNAs plays important roles in elucidating the biological function and biogenesis of miRNAs in living cells. However, because of their sequence similarity, low abundance, and small size, it is a great challenge to detect intracellular miRNAs, especially for those with much lower expression levels. To address this issue, we have developed an in cell signal amplification approach for monitoring down-regulated miRNAs in living cells based on biodegradable MnO nanosheet-mediated and target-triggered assembly of hairpins.

Metal-ion dependent DNAzyme recycling amplification for sensitive and homogeneous immuno-proximity binding assay of α-fetoprotein biomarker.

Based on target-induced immuno-proximity binding and metal ion-dependent DNAzyme recycling signal amplification, we describe the development of a homogeneous and sensitive fluorescent method for the detection of protein cancer biomarkers by using α-fetoprotein (AFP) as the model target. Two DNA strands with short complementary regions are conjugated to the AFP antibodies to prepare the recognition probes. The hybridization of the two DNAs in the absence of the AFP target molecules is inhibited due to the low melting temperature (T) of the complementary regions.

Dual-color encoded DNAzyme nanostructures for multiplexed detection of intracellular metal ions in living cells.

The detection of intracellular metal ions is of great importance in understanding metal homeostasis in cells and related diseases, and yet it remains a significant challenge to achieve this goal. Based on a new self-assembled and dual-color encoded DNAzyme nanostructure, we describe here an approach for multiplexed sensing of UO2(2+) and Pb(2+) in living cells. The fluorescently quenched nanoprobes can be prepared by simple thermal annealing of four ssDNAs containing the metal ion-dependent enzymatic and substrate sequences.

Multicolor-Encoded Reconfigurable DNA Nanostructures Enable Multiplexed Sensing of Intracellular MicroRNAs in Living Cells.

Despite the widespread utilization of gold nanoparticles and graphene for in vivo applications, complex steps for the preparation and functionalization of these nanomaterials are commonly required. In addition, the cytotoxicity of such materials is currently still under debate. In this work, by taking the significant advantages of DNA in terms of biocompatibility, nontoxicity, and controllability as building blocks for DNA nanostructures, we describe the construction of a reconfigurable, multicolor-encoded DNA nanostructure for multiplexed monitoring of intracellular microRNAs (miRNAs) in living cells.

RNA responsive and catalytic self-assembly of DNA nanostructures for highly sensitive fluorescence detection of microRNA from cancer cells.

Catalytic self-assembly of DNA nanostructures triggered by microRNA 21 (miR-21) is achieved through isothermal toe-hold strand displacement reactions. The miR-21 is autonomously recycled during the self-assembly process, which makes the generation of the DNA nanostructures proceed in a catalytic fashion. The miR-21-triggered self-assembly of DNA nanostructures can also serve as a remarkable signal amplification platform to achieve ultrasensitive detection of miR-21 from as low as 10 MCF-7 human breast cancer cells.

Click chemistry-mediated catalytic hairpin self-assembly for amplified and sensitive fluorescence detection of Cu(2+) in human serum.

Chemically reduced Cu(2+) triggers the ligation of alkynyl- and azido-modified DNA via click chemistry. Subsequently, the ligated DNA initiates cyclic assembly of two fluorescently quenched hairpin DNAs and generates significantly amplified fluorescence signals for highly sensitive detection of Cu(2+) in human serum samples.